Prebiotics in
Infant Nutrition
edited by:
SHARON DONOVAN, PhD
university of illinois—urbana, illinois
GLENN GIBSON, PhD
the university of reading—reading, united kingdom
DAVID NEWBURG, PhD
massachusetts general hospital—boston, massachusetts
Table of Contents
INTRODUCTION ...............................................................................................................................5
DEVELOPMENT OF INTESTINAL MICROBIOTA IN INFANTS ................................................6
COLONIZATION OF THE GI TRACT....................................................................................................6
BENEFICIAL FUNCTIONS OF INTESTINAL MICROBIOTA .................................................................8
FIGURE 1: BENEFICIAL FUNCTIONS OF INTESTINAL MICROBIOTA IN INFANTS ............................8
Metabolic effects ...........................................................................................................................8
Trophic effects ................................................................................................................................9
Protective effects ...........................................................................................................................9
DIFFERENCES BETWEEN MICROBIOTA OF BREASTFED AND FORMULA-FED INFANTS................11
BIOACTIVE COMPONENTS IN HUMAN MILK .........................................................................13
HUMAN MILK OLIGOSACCHARIDES ...............................................................................................14
HMOS influence GI function and immune protection .................................................................14
FIGURE 2: BENEFICIAL EFFECTS OF HUMAN MILK OLIGOSACCHARIDES
ON NEONATAL INTESTINAL DEVELOPMENT ...................................................................................15
USE OF PREBIOTIC CARBOHYDRATES IN INFANT FORMULAS ..........................................16
PREBIOTICS EVALUATED IN INFANT FEEDING ................................................................................16
TABLE 1: SELECT CHARACTERISTICS OF POTENTIAL PREBIOTIC
CARBOHYDRATES AND HUMAN MILK OLIGOSACCHARIDES ........................................................17
PREBIOTIC USE IN COMMERCIAL PRODUCTS .................................................................................18
EVIDENCE SUPPORTING USE OF PREBIOTICS IN INFANT FEEDING.................................19
EFFECTS OF PREBIOTICS ON GI MICROBIOTA .................................................................................19
Composition ..................................................................................................................................19
FIGURE 3: POTENTIAL BENEFICIAL EFFECTS OF PREBIOTICS ON INFANT HEALTH ........................20
GI health ........................................................................................................................................21
EFFECTS OF PREBIOTICS ON THE DIGESTIVE SYSTEM ....................................................................22
Laxation .........................................................................................................................................22
FIGURE 4: STOOL CONSISTENCY OF INFANTS FED PREBIOTIC-SUPPLEMENTED
FORMULA COMPARED TO INFANTS FED HUMAN MILK OR CONTROL FORMULA .......................22
Mineral absorption ........................................................................................................................23
EFFECTS OF PREBIOTICS ON IMMUNITY ....................................................................................................... 24
Biomarkers .....................................................................................................................................24
Incidence of allergy .......................................................................................................................25
Incidence of disease .......................................................................................................................25
CONCLUSION ....................................................................................................................................26
REFERENCES ......................................................................................................................................28
Executive Summary
Human milk fulfills many nutritive and immunoprotective functions for the neonate, especially
within the neonatal gastrointestinal (GI) tract. Although it is not possible to produce infant formula
that exactly replicates the composition of human milk, one goal of infant formula manufacturers is
to offer products that can simulate the composition and biological effects of human milk as closely
as possible. A very important area is the effect of human milk on content, composition and activity
of the gut microbiota.
Normal development and function of the GI tract critically depends on the presence of complex
microbiota, which perform numerous metabolic, growth-promoting, and protective roles. The
composition of the GI microbiota is influenced by ingestion of food substances that enhance
or inhibit growth of selected genera/species in the intestinal tract, mainly the colon.
Infants fed human milk have a predominance of bifidobacteria, a genus known to have healthpromoting functions, in their lower GI tract. The GI microbiota of breastfed infants also contain
low populations of potentially pathogenic groups like clostridia. The gut microbiota of formula-fed
infants, in contrast, are more diverse than those of breastfed infants, with similar or lower levels of
bifidobacteria, but greater diversity and higher levels of potential pathogens and commensal bacteria.
No single genus is thought to predominate in the formula-fed infant gut. Clinical and epidemiological
investigations have demonstrated that breastfed infants have lower rates of intestinal infection than
formula-fed counterparts, leading to the hypothesis that their decreased rates of infection compared
to formula-fed infants may be related in part to the higher concentration of bifidobacteria, which are
known to be inhibitory to pathogens. Differences in GI microbiota between breastfed and formulafed infants also result in different stool and laxation characteristics. Breastfed infants typically have
larger, softer, and more frequent stools than those who are formula-fed.
Although the microbiota present in the formula-fed infant may not necessarily be harmful,
it is desirable for infant formula to more closely match the functional activities of breast milk,
and investigators are currently seeking ways to promote the establishment of a similar intestinal
environment in formula-fed infants. Efforts to improve the performance of infant formula include
investigating the use of novel ingredients that have the potential to shift the GI microbiota of
formula-fed infants to more closely resemble those of breastfed infants.
Human milk oligosaccharides (HMOS) are a structurally diverse and highly variable group of bioactive
carbohydrates. HMOS resist digestion by enzymes in the human small intestine (including those
supplied from the pancreas) and thereby reach the lower GI tract largely intact. HMOS have several
key beneficial effects on the development of the neonatal intestine, including promotion of intestinal
colonization by beneficial microbes (e.g., bifidobacteria), stimulation of intestinal adaptation to
the extrauterine environment, compensation for developmental immaturity of the intestine, and
protection against colonization by pathogenic bacteria.
Infant formula manufacturers have evaluated numerous ingredients as part of continuing efforts to
more closely approximate the nutritional and functional properties of human milk, including those
of HMOS. Numerous studies have suggested that prebiotics can be used to alter the gut microbiota
of formula-fed infants to more closely simulate those of breastfed infants. A prebiotic has been
defined as a nondigestible food ingredient that brings about specific changes in the composition
and/or activity of the gastrointestinal microbiota which confer benefits upon host well-being and
health. Prebiotic ingredients are typically nondigestible oligosaccharides. Although it is impossible to
fully match the diverse and dynamic nature of oligosaccharides in human milk, a number of food-
2
grade oligosaccharides have demonstrated capacity to increase levels of selected beneficial bacteria
(principally bifidobacteria) and improve stool characteristics in formula-fed infants.
Commercial oligosaccharides studied for use in infant formula include galactooligosaccharides (GOS),
polydextrose (PDX), lactulose (LOS), inulin, and fructooligosaccharides (FOS), as well as combinations
of these products. The effects of prebiotics on infant growth, GI microbiota, stool characteristics,
mineral absorption and biomarkers of immune function have been assessed.
Current clinical evidence indicates that infant formulas supplemented with prebiotic carbohydrates
support normal growth and that prebiotics are capable of mimicking some of the effects of HMOS,
particularly in the areas of intestinal microbiota and stool characteristics. Clinical studies in infants
have shown that ingestion of GOS, LOS, inulin, and/or FOS can result in positive effects on the
composition of the GI microbiota. However, not all clinical trials have demonstrated a positive
effect of prebiotics on gut levels of beneficial bacteria. This may be due to differing methodologies
used to ascertain effects. As such, more studies are needed to establish a direct relationship between
alterations of infant GI microbiota and improved measures of GI health. Some studies have assessed
the effect of prebiotics on stool pH and short-chain fatty acid (SCFA) levels as a measure of the
health of the GI tract of infants. An additional goal of prebiotic supplementation is to produce
stool characteristics in formula-fed infants similar to those observed in breastfed infants. Studies
in infants have shown beneficial effects of prebiotic ingredients on stool characteristics.
Much of the evidence for the beneficial effects of GOS, PDX, LOS, or FOS on mineral absorption is
based on animal studies, which have shown that the absorption of calcium, magnesium, potassium,
and phosphorus was increased following dietary prebiotic supplementation. Although mineral
absorption in humans is positively affected with prebiotic supplementation, research has been
largely conducted in adolescents and postmenopausal women. Few clinical studies have evaluated
the effect of prebiotics on mineral absorption in infants and young children. In order to define the
role of prebiotics on mineral absorption, additional studies are needed—including a determination
of the mechanism of effect.
Because prebiotics influence the composition and activity of the gastrointestinal microbiota, which
in turn have a major effect on the immune system (mainly the gut-associated lymphoid tissue, or
GALT), it is reasonable to expect prebiotics to indirectly affect the immune system. Indeed, data
from animal studies suggest that dietary supplementation with prebiotics can influence certain
markers of immunity and affect the incidence of allergy and disease. However, the critical association
between prebiotic supplementation and improved immunologic function in humans remains to be
fully defined. Few studies have been conducted in humans, and most of these have been in adults.
More studies are needed to understand whether prebiotic ingredients in infant formulas lead to
sustained positive effects on infant immunity.
In summary, trials conducted in experimental animals and humans have shown beneficial
effects of prebiotics on the composition of GI microbiota, measures of GI health and immunity,
laxation, and mineral absorption. Mead Johnson will continue to evaluate the evolving science
supporting the use of prebiotic ingredients in infant formula; however, further research is
needed in order to provide state-of-the-science nutrition to formula-fed infants.
3
Introduction
Breast milk is widely regarded as the gold standard for infant feeding, and numerous health
organizations endorse breastfeeding as the optimal form of nutrition for infants during the
first year of life. However, over 80% of US infants receive formula at some time during the first
12 months of life.1 For these infants, iron-fortified formula is recommended.2 One goal for infant
formula manufacturers is to offer formulas that simulate human milk as closely as possible.
The gastrointestinal (GI) microbiota play a critical role in supporting health. For many years,
scientists have known that the composition of GI microbiota is influenced by dietary substances
that enhance or inhibit the growth of selected microbiota in the intestine. Numerous studies have
suggested that the gut microbiota of formula-fed infants can be altered to be more like those of
breastfed infants through the addition of ingredients called prebiotics. A prebiotic is a nondigestible
food ingredient that brings about specific changes in the composition and/or activity of the
gastrointestinal microbiota which confer benefits upon host well-being and health.3 A number of
potential prebiotic ingredients such as galactooligosaccharides (GOS), polydextrose (PDX), lactulose
(LOS), inulin, and fructooligosaccharides (FOS) has been evaluated for use in infant formulas.
Sidebar: Distinguishing Prebiotics from Probiotics
Prebiotics are food ingredients (typically oligosaccharides) that are selectively fermented
by beneficial bacteria in the gut (such as Bifidobacterium), stimulating the growth and/or
activity of those bacteria and thereby contributing to host health and well-being. Prebiotics
are resistant to gastric acidity, enzymes and absorption. Their purpose in infant formula is to
stimulate the growth and colonization of naturally occurring beneficial bacteria.
Probiotics are bacteria that pass through the GI tract and have beneficial effects on the health
of the host. Probiotic bacteria typically have a history of safe consumption; examples include
Lactobacillus rhamnosus GG and Bifidobacterium animalis ssp. lactis. Their purpose in infant
formula is to substitute for naturally occurring beneficial bacteria and thereby influence the
mucosal immune system.
4
5
Development of Intestinal Microbiota in Infants
The mature GI tract contains a large quantity and diversity of microbiota that play a critical role
in protecting and promoting infant health. Normal development and functionality of the human
GI tract depend on the presence of complex gastrointestinal microbiota, as these microorganisms
perform numerous metabolic, growth-promoting, and protective roles. 4, 5
Colonization of the GI tract
The infant GI tract is essentially sterile prior to birth, but is immediately exposed to maternal and
environmental bacteria during delivery. The subsequent colonization process is influenced by the
genetics of the host5 and several environmental factors, including mode of delivery, gestational age,
hospitalization, antibiotic use by the mother or infant, and type of infant feeding.6 Studies using
standard microbiological methods have shown that during the first few days of life, infants’
intestinal microbiota are comprised almost entirely of enterobacteria and Gram-positive cocci,
obtained predominantly during the birth process.7-9 Studies suggest that these facultatively
anaerobic bacteria generate conditions that favor the subsequent establishment of strict anaerobes
such as bacteroides, bifidobacteria, and clostridia.10, 11
The method of delivery at birth is important in determining the initial composition of intestinal
microbiota since infants’ GI tracts are first exposed to microorganisms during the delivery process.
Infants born by the vaginal route acquire their initial intestinal microbiota from maternal vaginal
and fecal microbiota, and it is recognized that the newborn infant’s fecal flora closely resemble
those of the mother. Infants born by Caesarean section show a different pattern of GI microbiota
development, which may also be influenced by other factors such as reduced gestational age,
increased exposure to the hospital environment, and prophylactic use of antibiotics.12, 13 Generally,
development of the GI microbiota is delayed for up to several weeks in Caesarean-delivered infants,
who have slower acquisition of Bifidobacterium and Bacteroides.12, 13 In preterm infants, intestinal
colonization with bifidobacteria is delayed.14, 15 This has been attributed to several factors including
delayed oral feeding, parenteral feeding, aggressive antibiotic therapy16 and reduced gut maturation
due to early gestational age.17
Diet has a particularly crucial influence on the composition of intestinal bacteria, especially in the first
few months of life.18 The intestinal microbiota of breastfed and formula-fed infants are comparable
for the first 3 to 4 days of life, but substantial differences between the two groups develop over
time.19 Another shift in GI microbiota composition typically occurs around 4-6 months of age with
the introduction of non-milk foods and complex carbohydrates to the diet. As weaning progresses,
both intestinal function and fermentation capacity continue to mature; composition of the intestinal
microbiota approximates that of adults by the second year of life.20
6
The predominant microorganisms in the adult human GI tract are highly individualized and, once
established, remain relatively stable over time.21 It is widely believed that microorganisms introduced
to the neonatal GI tract have a higher likelihood of becoming established compared to introducing
the same microorganisms (e.g., as probiotics) into the adult GI tract. By adulthood, the intestinal
microbiota are extremely complex, with total numbers reaching as high as 1012 bacterial cells per
gram feces and representing as many as 1000 different species.22, 23
Sidebar: Host-Microbe Interactions at Mucosal Surfaces:
Key to Balancing Immune Tolerance vs. Inflammation
It has long been appreciated that mutually beneficial relationships between animals and
microbes are a dominant theme of life; however, the extent to which our commensal
microbiota affect human health has been realized only recently. Human cells are outnumbered
ten-fold by the microbiota resident in the human GI tract, and the collective microbial
genome (microbiome) contains at least 100 times as many genes as our own.24 Humans have
co-evolved with these microbiota, which provide genetic and metabolic attributes that
humans do not possess, including the ability to harvest otherwise inaccessible nutrients.5 In
reality, the intestinal microbiota have a metabolic activity equal to a virtual “organ within an
organ,”25 and humans could be viewed as “superorganisms whose metabolism represents an
amalgamation of microbial and human attributes.”24 Beneficial microorganisms also partially
protect against pathogens by competing for metabolites, producing antimicrobial peptides,
occupying epithelial/mucous niches, and preventing host-pathogen interactions; thus, it is
in humans’ interest to tolerate them.26, 27
At the same time, however, immunity to dangerous microorganisms has to be initiated.
There are several ways in which the immune system may discriminate between commensal
and pathogenic microorganisms.27-29 Immune cells could discriminate between the two via
engagement of different pattern-recognition receptors (PRRs). Second, intestinal epithelial
cells could “sense” the presence of noninvasive (commensal) versus invasive (pathogenic)
microorganisms and transmit this information to antigen-presenting cells (APCs), such as
dendritic cells. Finally, APCs could be programmed to perform different tasks according to
their tissue of origin, such that dendritic cells or macrophages that are resident in the gut
mucosa and regularly encounter commensal bacteria are tolerogenic, whereas cells that are
recruited to the gut in response to pathogenic insult activate the immune response.27
7
Beneficial functions of intestinal microbiota
The primary functions of intestinal microbiota can be categorized as metabolic, trophic and
protective (Figure 1). Metabolic functions include fermentation of some carbohydrates to produce
short-chain fatty acids (SCFAs) and other end products,4, 20 with subsequent effects on luminal pH,
synthesis of vitamins and absorption of minerals.30-35 Trophic effects of the intestinal microbiota
include stimulating the proliferation and differentiation of colonic epithelial cells and proliferation
of specialized immune cells.36-46 One key protective function of the intestinal microbiota is to prevent
adhesion and colonization by pathogens. 4, 47 The net effect of these functional properties of the
intestinal microbiota is to protect the infant from infection, stimulate developmental maturation
of the gastrointestinal digestive and immune systems and promote growth and development.
Figure 1: Beneficial Functions of Intestinal Microbiota in Infants
Functions of Intestinal Microbiota
Metabolic
• Fermentation to
SCFA and other
end-products
• Energy salvage
• Vitamin synthesis
• Mineral
absorption
Trophic
Protective
• Proliferation and
differentiation of
colonic epithelial
cells
• Barrier against colonization
by pathogenic bacteria
• Proliferation
of intraepithelial
lymphocytes
• Stimulate production of
substances that inhibit
pathogen adhesion
(IgA, mucins)
• Production of organic acids,
which lower intraluminal pH
• Competition for receptors
and nutrients
Adapted from references 4 and 25.
Metabolic effects
One important function of the intestinal microbiota is to transform complex dietary and host
substances, such as mucin, into simpler compounds that can be used as energy sources by
other microorganisms and/or the host. Most notable is fermentation of non-digestible dietary
carbohydrates. 4, 20 The primary end products of this metabolic activity are SCFAs (primarily acetate,
8
propionate, and butyrate). Other end products include carbon dioxide, methane, and hydrogen.
Many of the beneficial effects of dietary fiber are attributed to SCFAs.34 These compounds are readily
metabolized by the intestinal epithelium, enhance small bowel digestion and absorption, stimulate
sodium and water absorption in the colon, and affect the immune function of the gastrointestinal
tract. SCFAs lower the gut pH, creating a more acidic environment that favors the growth of certain
beneficial bacteria and inhibits the growth of gram-negative and other potential pathogens. 4, 20, 30
The SCFAs are also readily absorbed and metabolized within the liver and contribute calories to
the host. Because of these effects, SCFAs have been the focus of much research in GI health.31, 32, 35
Additional metabolic effects of the gut microbiota include stimulation of the absorption of calcium,
magnesium, and iron, and metabolism of other dietary ingredients.33
Trophic effects
Intestinal microbiota increase the surface area of the intestinal lining by modulating crypt-villus
structure, epithelial turnover, GI motility, and development of the GALT.36 These trophic effects appear
to be mediated by SCFAs, which are readily absorbed by intestinal mucosal cells and have been shown
to increase epithelial proliferation and mucosal growth in the intestinal tract.31, 41-43, 45 These and other
changes in the colonic epithelial cells and in immune tissues and lymphocytes work synergistically
to decrease the likelihood of microbial and antigen translocation across the gut epithelial lining.
The SCFAs, particularly butyrate, stimulate the secretion of glucagon-like peptide-2 (GLP-2).34, 38 GLP-2
is a pluripotent hormone, which may represent a common mediator of SCFA-associated intestinal
proliferation, motility and nutrient absorption.39
The presence of a complex microbial community is integral to many functional processes in the
human GI tract. For example, colonization of germ-free mice with Bacteroides thetaiotaomicron,
which is a normal constituent of both human and mouse intestinal microbiota, significantly altered
the expression of host genes related to nutrient absorption, mucosal barrier function, xenobiotic
metabolism, angiogenesis and maturation. 40 Others have shown that components of the intestinal
microbiota stimulate angiogenesis (vascularization),46 provide protection against epithelial cell
injury,44 and affect host fat storage.37 Because of ethical issues involved in human studies, much of
our understanding about how colonizing microorganisms affect infant GI development and function
has been derived from animal studies.
Protective effects
One of the key functions of the intestinal microbiota is to provide a natural barrier against
colonization and proliferation of opportunistic pathogens, thereby decreasing the risk of
intestinal infection and disease. 48 The ability of certain intestinal bacteria, such as bifidobacteria
and lactobacilli, to interfere with colonization by exogenous and possibly disease-causing
microorganisms is called “colonization resistance”.10 Selective stimulation of beneficial
genera in the infant GI tract at the expense of harmful bacteria may also be referred to as
“competitive exclusion.” It is widely reported that consumption of breast milk by infants
can promote the growth of beneficial bacteria such as Bifidobacterium spp16 and/or
reduce colonization by less beneficial or potentially harmful species, such as Escherichia
coli (E. coli) and Clostridium difficile (C. difficile). 49 This method of inhibiting colonization
9
by undesirable bacteria has been proposed as one protective mechanism that helps improve the
resistance of breastfed infants to infection.
Indigenous intestinal microbiota can inhibit the growth of potentially pathogenic exogenous
microorganisms in several ways, such as:
• Competing with pathogens for available nutrients;
• Producing organic acids, which lower intraluminal pH to a level that inhibits pathogen growth;
• Stimulating the production of substances (mucins and IgA) that inhibit pathogen adhesion
to the gut wall or block their sites of colonization30;
• Excreting antimicrobial factors such as peptides; and
• Compromising the virulence of potential pathogens by preventing attachment to epithelial
cells, competing for nutrients and producing bacteriocins4
Sidebar: Methodological Issues with Evaluating GI Microbiota
Advances in molecular biology have made it possible to more fully understand the effects of
diet on GI microbiota. Classical microbiology techniques attempt to recover specific bacterial
species using partially selective artificial media containing complex blends of nutrients,
growth factors, antibiotics, and other ingredients. Such methods have been reported to
recover as few as 10% of total fecal bacteria,50 partly due to the strictly anaerobic growth
environment of nearly all target microorganisms.11 However, these methods are now seen as
flawed because of a lack of media selectivity, an inability to recover non-culturable diversity
and the subjective nature of colony descriptions.
More recent non-culture-based methods that identify bacteria by specific ribosomal DNA
(rDNA) sequences have proven essential in gaining a more accurate representation of the
infant GI microbiota.51 Four main approaches have been used:
• The first involves amplification of rDNA target sequences of all bacteria in a stool
sample by polymerase chain reaction (PCR). Separation of the resulting bands
by denaturing gradient gel electrophoresis (DGGE) or temperature gradient gel
electrophoresis (TGGE) and sequence analysis make it possible to reveal the
biodiversity of the intestinal microbiota.11
• The terminal restriction fragment length polymorphism (T-RFLP) approach
involves amplifying and fluorescently labeling bacterial rDNA and then digesting
amplicons with restriction enzymes to generate terminal-restriction fragments
(T-RFs). The T-RFs are separated by gel electrophoresis and sequenced and the
sequences aligned with known bacterial sequences.52
• In addition, 16S rRNA sequences, species- and group-specific primers can also be
used to directly identify a microbe of interest by quantitative real time PCR (RTPCR).49
• Lastly, bacteria in stool samples can be identified by fluorescence in situ
hybridization (FISH) analysis using fluorescently-labeled, species-specific rDNA
primers.53
Differences between microbiota of breastfed and formula-fed infants
Both conventional and molecular microbiology techniques have shown that infants fed human milk
have a GI microbiota profile containing a predominance of bifidobacteria,54 which are known to have
health-promoting functions. Bifidobacteria are particularly predominant among infant GI microbiota,
consisting primarily of B. breve, B. longum, and B. infantis, along with an emerging but significant
contribution of other genera.51 DNA-based techniques have shown that Bifidobacterium species
appeared in the feces of newborns by 5 or 6 days postpartum in breastfed and formula-fed infants,
respectively.51 In general, results also suggest that the GI microbiota of breastfed infants contain
reduced populations of potentially pathogenic species such as C. difficile and E. coli. 49
In contrast, the gut microbiota of formula-fed infants are more diverse and similar to those
observed in adults, with lower levels of bifidobacteria55, but greater diversity and higher levels of
other potentially pathogenic groups, including Bacteroides, Clostridium, and Enterobacteriaceae.9,16, 56-59
While not all studies are in agreement, many have confirmed these overall patterns, including more
recent studies using reliable and accurate molecular-based methods.50, 60
Sidebar: What Makes Bacteria Harmful in the Body?
It is normal for the human GI tract to contain both beneficial and potentially harmful types
of microbes. Their effects on host health depend on the balance of these types of microbes,
the health of the host and numerous factors within the environment of the GI tract. When
beneficial bacteria are predominant, potentially pathogenic microbes are less likely to
multiply to levels at which they could cause illness.
Major mechanisms by which pathogenic bacteria can become harmful include attachment to
epithelial cells via specific receptors and/or secretion of toxins (e.g., Cholera toxin). Interaction
of bacteria with host cells triggers an immunological response in the host, including the
secretion of proinflammatory cytokines at the site of infection. When the intestinal barrier
function is impaired, bacteria may translocate across the intestine and cause septicemia.
Results from several studies have been compared to determine whether patterns of infant fecal
bacteria are reflective of feeding regimen.22 Fecal samples from breast- and formula-fed infants were
not different in overall levels of bifidobacteria or lactobacilli. However, higher levels of clostridia were
observed in the feces of formula-fed infants than breastfed infants. Fecal levels of enterobacteria and
streptococci also tended to be lower in breastfed infants. In a more recent study, RT-PCR techniques
were used to monitor the GI microbiota of exclusively breastfed and formula-fed infants. 49 All of the
infants were colonized by Bifidobacterium species, and mean values were comparable among the
breastfed and formula-fed groups. In contrast, the prevalence and counts of both C. difficile and E. coli
were significantly lower in breastfed infants.
Molecular methods have been particularly useful in evaluating beneficial microbiota such as
bifidobacteria at the genus and species levels, where classical microbiology can provide little
or no information.
10
11
Bioactive Components in Human Milk
This finding is of clinical relevance because pathogenic bacteria have been strongly
associated with disease in infants. For example, C. difficile is responsible for 20-40%
of cases of antibiotic-associated diarrhea61 and has been implicated in necrotizing
enterocolitis.62 There are extensive data demonstrating the protective role of the
intestinal microbiota against C. difficile. 48, 63-65 In fact, it is thought that the normal
flora suppress the activity of C. difficile and that this suppression becomes compromised
during antibiotic intake.
Differences in GI microbiota between breastfed and formula-fed infants also result in
varying stool and laxation characteristics. Breastfed infants have larger, softer, and more
frequent stools than formula-fed infants, which is attributed to differences in bacterial
cell mass, fecal pH, and other factors.
Although the microbiota present in the formula-fed infant may not necessarily be harmful,
it is highly desirable for infant formula to function more like breast milk, and investigators
are currently seeking ways to create a similar intestinal environment in formula-fed infants.
Efforts to improve the performance of infant formula include the use of novel ingredients
that have the potential to shift the GI microbiota of formula-fed infants to more closely
resemble that of breastfed infants.
Human milk is a complex physiological fluid; its components fulfill many nutritive, developmental,
and immunoprotective functions in infant nutrition and within the neonatal GI tract. Various
protein, lipid, and carbohydrate components of human milk are involved in stimulating growth and
maturation of cells in the GI tract, aiding in the establishment of the gastrointestinal microbiota,
enhancing mucosal barrier function, and stimulating development of the GALT, which functions
as a crucial interface between intestinal contents and the infant’s developing immune system.66
Bioactive components of human milk are thought to provide protection while the infant develops
greater immunological and developmental maturity.67 This concept is supported by clinical studies
demonstrating that the incidence and severity of infectious diseases are lower in breastfed infants
compared to formula-fed counterparts.55, 68-73
Proteins, lipids, carbohydrates, and other bioactive components in human milk provide essential
nutrients and energy; they also function to support growth and development in other ways.
Categories of bioactive compounds in human milk include immune factors (such as secretory
IgA, or sIgA), hormones, growth factors, proteins and peptides, lipids, carbohydrates, and other
components.55 It has been suggested that the bioactive substances in human milk represent an
evolutionary strategy for protecting infant health and promoting gastrointestinal development
during the vulnerable newborn period.74 Beneficial functions associated with bioactive compounds
in human milk include:
• Compensating for developmental immaturity;
• Protecting against infection by potential pathogens;
• Aiding intestinal adaptation to extrauterine life;
• Reducing inflammation of the GI tract;
• Protecting bioactive substances from digestion;
• Aiding the establishment of a beneficial microbial population.74
A review of the literature describing bioactive components typically identified in human milk offers
a number of insights into how they might influence neonatal GI development. For example, the
whey protein fraction of human milk contains high concentrations of hormones, growth factors
and numerous antimicrobial proteins.66 The nutritional, developmental and protective contributions
of human milk whey proteins have been investigated for several years. Scientists have only recently
begun evaluating the contributions of oligosaccharide components in human milk. The importance
of HMOS is evidenced by their relative prominence; HMOS constitute the third-largest component
of human milk, following lactose and lipid, and are thought to play significant developmental and
protective roles.75
12
13
Human milk oligosaccharides
Human milk oligosaccharides are a structurally diverse and highly variable group of bioactive
carbohydrates. HMOS are comprised of a mixture of oligosaccharides that differ in molecular
weight and structure, constituent sugar or sugar derivatives, and charge.76 Structurally, HMOS
are composed of combinations of five monosaccharide building blocks: D-glucose, D-galactose,
N-acetylglucosamine, L-fucose and sialic acid. Some of these monosaccharides are also found in
the milk of other mammalian species.77 HMOS resist digestion by enzymes in the stomach and small
intestine and reach the lower GI tract largely intact. They also can be absorbed into the circulatory
system and transported to other sites, such as the urinary tract, where they are thought to function
systemically by blocking pathogen adhesion77 as many of the structures present in HMOS resemble
typical pathogen binding sites (i.e., the HMOS thereby act in a ‘decoy’ manner).
Variability occurs in both the concentration and composition of HMOS during the normal course
of lactation.78 Unbound HMOS are found in mature milk at levels ranging from 5-10 g/L and are
present at higher levels in colostrum.77 They have been estimated to vary in size from 3 to 32 sugars.75
Current analytical methods have detected approximately 200 distinct molecular species of HMOS,79
though this may represent only a small fraction of their true structural diversity.80 Further, HMOS
concentrations vary considerably among women and during lactation. As a result of this complexity,
it is not possible to replicate the HMOS content or composition of human milk in infant formula.
HMOS influence GI function and immune protection
A significant proportion of HMOS pass undigested into the lower gastrointestinal tract,81 where
they can be selectively metabolized by beneficial microorganisms such as the bifidobacteria.82 The
bifidogenic capacity of HMOS is frequently cited as an important reason why the gastrointestinal
microbiota of breastfed infants contain proportionally more bifidobacteria than those of formulafed infants.54 In fact, the original “bifidus factor” is human breast milk. Human milk oligosaccharides
have several key beneficial effects on the development of the neonatal intestine (Figure 2). 66
Figure 2: Beneficial Effects of Human Milk Oligosaccharides
on Neonatal Intestinal Development
Promote establishment
of beneficial
microbiota
Protect against
infection
Oligosaccharides
Help compensate
for developmental
immaturity of the
intestine
Promote intestinal
adaptation to
extrauterine
environment
Adapted from reference 66.
Oligosaccharides are constituents of the innate immune system in human milk that provides
breastfed infants with protection from potential pathogens. The significant immunological
protection afforded to infants by human milk has been attributed, in part, to the presence of
α(1-2)-linked fucosylated oligosaccharides.75 This is supported by observations that fucosylated
oligosaccharides are capable of preventing diarrheal illness through diverse mechanisms, including:
1) inhibition of E. coli stable toxin activity by binding and blocking access to target receptors; 2)
prevention of Campylobacter adhesion to intestinal cells; and 3) competitive inhibition of the binding
of Norovirus to the intestinal epithelium.83
In contrast to human milk, cow’s milk contains only trace amounts of oligosaccharides. Infant
formulas made from cow’s milk are essentially devoid of oligosaccharides, unless they are provided
exogenously. Although it is not possible to add oligosaccharides that are structurally identical
to HMOS to infant formula, researchers have evaluated the feasibility and desirability of adding
food-grade prebiotic ingredients to partially mimic the functional properties of HMOS. The
intended benefit of such prebiotic ingredients in infant nutrition is to bring the biological
responses of infants to formulas into closer alignment with breast milk, particularly with
regard to development of positive intestinal microbial populations and associated effects
on immunity and stooling patterns.
14
15
Use of Prebiotic Carbohydrates in Infant Formulas
Given the many benefits purported to be associated with the intestinal microbiota present
in breastfed infants, it is not surprising that researchers are interested in determining ways to
promote similar microbiota in formula-fed infants. The addition of prebiotics to infant formula
has the potential to mimic some of the beneficial effects of HMOS in formula-fed infants. One
goal of using prebiotics in the diet is to modify the intestinal microbiota such that their beneficial
activities are enhanced and detrimental activities suppressed. To be effective, prebiotics must be
resistant to digestion until they are specifically fermented by the intestinal microbiota. Also, by
definition, prebiotic substances must have health-promoting properties.3 Postulated benefits of
prebiotic ingredients for infants include increased production of SCFAs, stimulation of growth
and/or activity of lactic acid bacteria, improved stool characteristics (laxation) and improved
bioavailability of minerals.84
Infant formula manufacturers have evaluated numerous ingredients as part of their continuing effort
to more closely approximate the nutritional and functional properties of human milk. Although it is
impossible to completely match the diverse and dynamic nature of HMOS, a number of food-grade
oligosaccharides have demonstrated capacity to increase levels of select beneficial bacteria (typically
bifidobacteria) and improve stool characteristics in formula-fed infants.19
Prebiotics evaluated in infant feeding
Several food-grade oligosaccharides have been evaluated for use as prebiotics in infant
formula, including galactooligosaccharides (GOS), polydextrose (PDX), lactulose (LOS), inulin,
and fructooligosaccharides (FOS). Combinations of these products have also been evaluated.
Oligosaccharide ingredients may be commercially produced by bacterial synthesis, enzymatic
synthesis or extraction from natural sources.85 Dietary components most commonly investigated
as potential prebiotics are nondigestible carbohydrates, primarily oligosaccharides, consisting of
2 to 20 saccharide units.86 Dietary oligosaccharides are present in fruits such as bananas and in
vegetables such as asparagus, leeks, and artichokes. Many naturally-occurring oligosaccharides
are relatively resistant to digestion by intestinal enzymes and subsequently fermented by bacteria
in the colon. There has been considerable interest in supplementing foods with prebiotic ingredients
because the levels of naturally-occurring oligosaccharides in diets are considered inadequate to
produce substantial effects on health.87
Selected characteristics of these prebiotics and human milk oligosaccharides are shown in Table 1.
Table 1: Select Characteristics of Potential Prebiotic Carbohydrates and
Human Milk Oligosaccharides
Prebiotic
Carbohydrate
Abbreviation
Chemical
Structure*
Degree of
Polymerization†
3-30
12 Average
Galactooligosaccharides
GOS
Polydextrose
PDX
Lactulose
LOS
α-D-Glu-(1→4)[β-D-Gal-(1→6)]n
Random, highly
branched glucose
polymer + sorbitol
4-O-β-D-Gal-D-Fru
IN
Glu-[β-Fru-(2→1)]n
Inulin
Fructooligosaccharides
Human Milk
Oligosaccharides
FOS
HMOS
Glu-[β-Fru-(2→1)]n
Fru-[β-Fru-(2→1)]n
Complex and
highly variable
2-8
2-4 Average
2 (Disaccharide)
3-60
10-12 Average
2-8
4 Average
3-32 (Estimated)
* Abbreviations: Glu=glucose; Gal=galactose; Fru=fructose; O=oxygen.
†
Degree of polymerization=number of repeating carbohydrate units (represented by n)
Given that human milk contains a wide variety of oligosaccharides, it has been postulated that
the benefits of HMOS on infant health are more likely associated with a mixture of oligosaccharide
structures rather than a single oligosaccharide.88 A combination of GOS and PDX was shown to result
in a range of oligosaccharide chain lengths within the range reported for HMOS.75 Blends of GOS and
FOS have been widely studied and found to be effective in stimulating the growth of bifidobacteria
and promoting the establishment of intestinal microbiota similar to that of breastfed infants.89
In vitro studies measuring carbohydrate utilization patterns and the production of short-chain
fatty acids and gas by infant fecal bacteria have shown that larger, more complex carbohydrates,
such as PDX and inulin, are fermented more slowly and less completely than short-chain
materials such as LOS, FOS, and GOS.90 Thus, a combination of specific long-chain and short-chain
carbohydrates may allow for slower fermentation by fecal bacteria, a process that may translate
into a more sustained effect during gastrointestinal transit. This is a desirable trait as the distal (left)
side of the large intestine has low saccharolytic activity compared to more proximal areas and is also
more frequently affected by intestinal disorders. Further, in vitro data suggest that blends of prebiotic
carbohydrates would be more likely to stimulate fermentation by a broader array of gastrointestinal
bacteria, resulting in greater SCFA production and reduced pH—both conditions that are considered
unfavorable for pathogens.
“The addition of certain mixtures
of oligosaccharides in infant formula
bring infant formula, the second choice
infant feeding, one step closer to the
gold standard, breast-feeding.”19
16
17
Prebiotic use in commercial products
There appears to be broad support for the use of prebiotics in infant formulas in many countries,
judging by the commercial availability of prebiotic-supplemented formulas. Prebiotic carbohydrates
have been added to infant formulas in Japan for over 20 years, and 90% of infant formulas in Japan
are purported to contain prebiotics.88 Within the past decade, prebiotics have been introduced into
formulas in Europe as well.88 Prebiotic-supplemented infant formula is a relatively new concept in
the United States. In recent years, however, there has been an increase in the number of foods
available for children and adults containing prebiotics, including yogurt and cereals.
Evidence Supporting Use of Prebiotics in
Infant Feeding
More than 30 research articles have described results of infant clinical trials with prebiotics.
A key finding of a majority of trials is that infants consuming prebiotic-supplemented formulas
have significantly increased levels of bifidobacteria compared to those consuming control
formulas. The results have been equivocal, however, as a bifidogenic response has not always
been demonstrated, even in studies using the most reliable (molecular-based) methodologies.91
Studies that evaluated growth have shown that formulas containing prebiotics support growth
comparable to that observed in infants fed control formulas. Another consistent finding with
prebiotic supplementation is beneficial changes in stooling patterns (i.e., increased frequency
and softness).
A study in term infants (n=226) found that infants fed formula supplemented with a mixture of
prebiotics had similar growth up to 120 days of age as those fed unsupplemented (control) formula.92
Infants in one study group received formula supplemented with PDX and GOS (50:50 ratio) at a
level of 4 g/L; infants in a second study group received formula with 8 g/L of PDX, GOS, and LOS
(50:33:17 ratio). There were no significant differences in weight or length growth rate between the
supplemented and control formula groups from 14 to 30, 60, 90 or 120 days. Studies with other
prebiotics and prebiotic blends have likewise noted no differences in growth between infants fed
supplemented or unsupplemented formulas.93-95
Effects of prebiotics on GI microbiota
Clinical studies in infants suggest that prebiotics have positive effects on measures of
gastrointestinal health (Figure 3). Individual prebiotic ingredients and blends of prebiotic
ingredients have been evaluated. More studies in infants are needed to establish a relationship
between alterations in GI microbiota and improved measures of GI health.
Composition
A few studies have evaluated the effectiveness of individual prebiotics on gut microbial composition.
In one study with 271 term infants, subjects who consumed formula with GOS added (2.4 g/L) had
significantly increased fecal levels of bifidobacteria and lactobacilli after 3 months and 6 months
of feeding.96 Importantly, there were no significant differences in levels of bifidobacteria or
lactobacilli between GOS-supplemented infants and those in a breastfed control group. Another
study in preterm infants evaluated the effect of inulin on microbiota.97 In this study, 56 preterm
infants received formula with or without inulin (4 g/L) for 7 days. Fecal bacterial counts showed
that the microbiota of supplemented infants were more heavily colonized with bifidobacteria
and had fewer potentially pathogenic bacteria (E. coli and Enterococcus) compared to infants
fed unsupplemented control formula.
18
19
Figure 3: Potential Beneficial Effects of Prebiotics on Infant Health
Prebiotics
Softer stools
Not all clinical trials have demonstrated a positive effect of prebiotics on gut levels of beneficial
bacteria, however. In one study of 72 infants supplemented with FOS at 1.5 g/L or 3.0 g/L for 7 days,
only mild prebiotic effects were reported.108 In another study with 76 infants, supplementation with
FOS at 2 g/L for 13 weeks did not result in significant increases in fecal levels of bifidobacteria and
lactobacilli compared to control formula.91
Sidebar: Understanding Differences in Study Outcomes
Increased Beneficial Bacteria
Positive effect on
local GI immunity
Positive effect on
general immunity
Increased SCFA
production
Increased gut
maturation
Decreased
pathogenic
bacteria
Infant feeding studies with prebiotic ingredients are not always consistent in their results.
Differences in study design can have important effects on the outcomes observed, and such
differences should be considered when comparing study results. Key elements of study
design that can impact outcomes include:
Lower pH
Differences in one or more of these study parameters can lead to outcomes that appear
to be contradictory. In fact, gastrointestinal microbiota exist in a complex environment
that is affected by many different variables. Also, it is important to keep in mind that genetic
differences among infants are likely to also influence initial colonization and response
to prebiotics.
Improved nutrient
absorption
A larger number of studies in infants has evaluated the effects of blends of 90% (w/v) GOS and
10% (w/v) FOS or inulin.98 In many of the studies, FOS and inulin are used interchangeably, as inulin
is long-chain FOS. The majority of these studies showed a positive correlation between prebiotic
supplementation and enhanced populations of bifidobacteria18, 94, 99-104 and lactobacilli.100, 103, 105 Many
of the studies also demonstrated decreased levels of potentially pathogenic bacteria with prebiotic
supplementation. In a few studies, the investigators used techniques that allowed analysis of the
microbial populations at a species-specific level, which demonstrated that infants fed the GOS- and
FOS-supplemented formulas had increased total numbers of beneficial bacteria (both Lactobacillus
and Bifidobacterium) as well as a species-specific profile that more closely resembled that of
breastfed infants.101, 103, 105 These clinical trials suggest that combinations of prebiotics may be helpful
in altering the composition of gut microbiota to be more like that of breastfed infants.
A clinical trial of 90 infants examined the effects of a blend of 90% (w/v) GOS and 10% (w/v) FOS
on bacterial growth.106, 107 Infants were randomized to receive control formula or formula
supplemented with either 4 g/L or 8 g/L of the GOS:FOS blend for 4 weeks. Bifidobacteria counts
were significantly increased in both supplemented groups compared to the control group at the
end of the treatment period; this effect was dose-dependent and significantly different between
the supplemented groups (P<0.01). Lactobacilli were also significantly increased in the supplemented
infants compared to the control group, but there was no statistically significant difference between
the two supplemented groups.
20
• Length of gestation and postnatal age of infants being evaluated
• Mode of infants’ delivery
• Duration of supplementation
• Type and level of prebiotic ingredients used
• Methods for measuring study outcomes, especially gut microflora changes
• Use of medications or supplemental feeds
GI health
In addition to evaluating composition of the GI microbiota, some studies have assessed the pH and
SCFA levels of infant stools as a determinant of GI tract health. Prebiotic supplementation resulted
in lower stool pH18, 53, 100 and increased levels of fecal SCFAs.18, 53 In one of these studies,18 53 infants
were randomly assigned to receive either control formula or formula supplemented with GOS
and FOS (9:1 ratio, 8 g/L) for 6 weeks. Stool pH of the supplemented group was significantly lower
than that of the control group (pH of 5.7 vs. 6.3; P<0.001) and did not differ from that of a breastfed
reference group. Further, the SCFA profiles of the supplemented and breastfed infants were similar,
but the SCFA profiles of supplemented and control infants differed.
Effects of prebiotics on measures of barrier function in the GI tract are difficult to evaluate in
infant studies. However, prebiotic oligosaccharides have been shown to block pathogen adhesion
in vitro. Shoaf and coworkers evaluated the capacity of a range of commercially available prebiotic
oligosaccharides to block the attachment of E. coli to certain human-derived cell lines. Purified GOS
exhibited the greatest activity of the oligosaccharides evaluated, inhibiting E. coli adherence to
intestinal epithelial cells by up to 70%.109
21
Effects of prebiotics on the digestive system
Mineral absorption
Studies in animals and humans have demonstrated that supplementation with prebiotics
has positive effects on the digestive system, including improved laxation and increased
mineral absorption.33
Laxation
An overall goal of prebiotic supplementation is to produce stool characteristics in formula-fed infants
similar to those observed in breastfed infants. A number of studies in infants have shown beneficial
effects of prebiotic ingredients on stool characteristics. Positive results have been observed with GOS
(softer stool consistency)96 and FOS (more frequent stools and softer stools).110
Prebiotic blends have also demonstrated positive results. In a study of more than 200 infants,
formulas were supplemented with either 4 g/L of PDX and GOS (50:50 ratio) or 8 g/L of PDX,
GOS and LOS (50:33:17 ratio) and fed through 120 days of age.92 Prebiotic supplementation with
either amount resulted in a stool consistency that was intermediate between that of infants
fed unsupplemented formula92 and that of infants in another study who were fed breast milk
(Data on file, 2005) (Figure 4).
Positive effects on infant stool consistency and frequency have also been observed with prebiotic
blends composed of 90% (w/v) GOS and 10% (w/v) FOS. These effects include fewer symptoms of
constipation111; increased stool frequency106, 112, 113 and softer stool consistency.106, 112, 114 Several studies
showed that infants who received the prebiotic blend had stool consistency similar to that of
breastfed infants.106, 112, 114
Figure 4: Stool Consistency of Infants Fed Prebiotic-supplemented Formula
Compared to Infants Fed Human Milk or Control Formula
100
% of Infants with Loose
or Watery Stools
90
Researchers are interested in the potential for prebiotic ingredients to increase absorption
of minerals. Even though most minerals are absorbed primarily in the small intestine, it is
thought that a lower pH in the colon, resulting from by-products of bacterial fermentation
(including SCFA), increases the solubility of some minerals thereby potentially increasing
their absorption from the colon.33, 115
Studies evaluating the effect of prebiotics on mineral absorption primarily have been conducted
in animals. Most of the evidence for the beneficial effects of GOS, PDX, LOS, or FOS on mineral
absorption is based on animal studies; these have shown that the absorption of calcium, magnesium,
potassium and phosphorus was increased after prebiotic diet supplementation.116-124 Animal studies
also suggest that supplementation with GOS and LOS may increase intestinal absorption of calcium
and magnesium; results of a study conducted with PDX showed an increase in calcium absorption
and bone mineralization.125 The applicability of these studies to infants and children has not yet
been determined.
Although studies of mineral absorption in humans have shown positive results with prebiotic
supplementation, they have been conducted primarily in adolescents and postmenopausal
women. One study with children in India suggested a positive effect of GOS on iron absorption,
but confounding study variables make it difficult to conclude that the effect was due to the
supplement.126 In a clinical trial with 30 preterm infants, urine calcium concentrations tended
to be higher in infants fed a GOS- and FOS-supplemented formula compared to a placebosupplemented formula, despite equal dietary calcium intake.127
In summary, few clinical studies have evaluated the effect of prebiotics on mineral absorption
in infants and young children, and it is difficult to extrapolate results from studies in adolescents
and adults to infants due to differences in their GI systems. Further, because prebiotics may also
decrease GI transit time, there is a possibility that nutrient absorption may be diminished if the
prebiotic level is too high. In order to define the role of prebiotics in mineral absorption, additional
studies are needed to address issues such as optimal intake, duration of supplementation and the
impact of dietary calcium intake.
80
Human Milk1
70
60
50
40
Prebiotics (8 g/L)
Prebiotics (4 g/L)
Control
30
20
10
0
30
60
90
120
Age (Days)
Formula data are from Zeigler et al. (92); Human milk data are from Mead Johnson (Data on file, 2005).
1
22
23
Effects of prebiotics on immunity
The gut microbiota of breastfed infants are dominated by bifidobacteria, and it has long been
suggested that they play a role in enhancing infant resistance to infection.106 Analyses of fecal
bacteria using species-specific probes have demonstrated that infants fed prebiotics develop
microbiota more similar to those of breastfed infants.101, 105 Because prebiotics influence the
composition and activity of the gastrointestinal microbiota, and because the microbiota are
known to have a major effect on the immune system, it is reasonable to expect that prebiotics
can be used indirectly to modulate the immune system. Data from animal studies suggest that
dietary supplementation with prebiotics can influence certain markers of immunity and affect
the incidence of allergy and disease. However, the critical association between prebiotic
supplementation and improved immunologic function in humans remains to be defined.
The effects on immunity of ingesting GOS, LOS, PDX, or blends thereof have not hitherto been
examined in infants or children. There is a need for further studies in humans to evaluate the effect
of prebiotic ingredients on the gut immune response and subsequent effects for overall health.
Biomarkers
Most of the evidence for the beneficial effects of prebiotics on immunity is based on animal
studies. These studies have evaluated the effectiveness of prebiotic ingredients in reducing
intestinal infection by decreasing the translocation of bacteria (particularly pathogens) from
the intestine to the liver and mesenteric lymph nodes. Various markers of immune function have
been used to measure this effect, including concentrations of endotoxin, TNF-α, interleukin-1,
interleukin-10, and IgA; levels of pathogenic bacteria; inflammation; and incidence of intestinal
infections. In vitro and animal studies have suggested that the addition of complex fermentable
carbohydrates to the diet can modulate the type and function of cells from different regions of the
gut-associated lymphoid tissue,128-130 increase immunoglobulin production in the small intestine and
cecal mucosa131, and alter the profile of inflammatory cytokines in plasma and intestinal cells.132-134
Few studies in infants have specifically evaluated the effect of prebiotic supplements on markers
of immune function. One study with 57 infants evaluated the effect of formula supplemented with
FOS and GOS (6 g/L for 32 weeks) on fecal levels of IgA.135 Supplemented infants displayed a trend
toward higher fecal sIgA compared to unsupplemented infants, but this difference reached statistical
significance only at week 16.
24
Incidence of allergy
Exclusive breastfeeding is strongly recommended for newborn infants with a family history of
allergy, as breastfeeding reduces the likelihood that the infant will develop atopic disease. One
study in infants directly evaluated whether infant formula supplementation with prebiotics could
replicate the protective effect of breastfeeding. In this study, 259 infants at risk for atopic disease
were randomized to receive either control formula or formula supplemented with a blend of GOS
and FOS (9:1 ratio; 8 g/L). 136 Fecal microbiota were analyzed in a subgroup of 98 infants. Levels of
bifidobacteria were significantly higher among infants who received the prebiotics compared to
control infants; levels of lactobacilli did not differ between the groups. Over the 6-month course of
the study, fewer infants in the supplemented group developed atopic dermatitis compared to the
control group, suggesting that prebiotics may modulate postnatal immune development in part by
altering the GI microbiota; there may be other mechanisms as well. This study supports a potentially
positive role for prebiotics in managing symptoms of allergy during infancy, but additional studies
are still needed.
Incidence of disease
Breastfed infants are often reported to experience a lower incidence of disease than formula-fed
infants.137 This effect is attributed to multiple factors associated with breastfeeding, including the
presence of maternal antibodies138 and oligosaccharides in human milk.75 Although scientists are
hopeful that the use of prebiotics and other ingredients can affect markers of disease in infants, few
studies have directly evaluated the relationship between prebiotic supplementation and incidence
of disease or reduction of risk. One clinical study in children suggested that supplementation with
FOS (2 g/d) for 21 days could improve their immunological status, based on a lower incidence of fever,
vomiting, and diarrhea in those taking the supplement.139 Another study, conducted among breastfed
infants living in a community near Lima, Peru with a high prevalence of GI and other infections, found
that feeding infant cereal supplemented with oligofructose at 0.55 g/15 g cereal for 6 months was
not associated with change in incidence of diarrhea, use of health care resources, or response to a flu
vaccination.140 A high prevalence of breastfeeding in the study population was thought to contribute
to the lack of effect with prebiotics observed. Further studies in infants and children are needed to
clarify these findings.
25
Conclusion
Oligosaccharides in human milk are constituents of an innate immune system that provides
breastfed infants with protection from potential pathogens. Oligosaccharides are prominent in
human milk, as the third largest component, and are thought to play significant developmental
and protective roles.75 Human milk oligosaccharides are active in the development of intestinal
microbiota in breastfed infants, leading to increased gut levels of beneficial bacteria, particularly
bifidobacteria, and reduced levels of potentially pathogenic bacteria. The bifidogenic potential of
HMOS is thought to be one important reason why the gastrointestinal tract of breastfed infants
contains proportionally more bifidobacteria than that of formula-fed infants.54
Because of their complexity and variability, it is not possible to duplicate HMOS for use in infant
formula. As an alternative, the safety and efficacy of naturally occurring and manufactured
oligosaccharides in infant feeding have been evaluated. Findings from animal and human studies
suggest that prebiotic ingredients can have some effects similar to those of HMOS in the GI tract.
Postulated benefits in infants include increased production of SCFAs, inhibition of growth and/or
activity of pathogenic bacteria, stimulation of growth and/or activity of beneficial bacteria, improved
stool characteristics, and improved bioavailability of minerals.84 Studies have demonstrated that
infants fed prebiotics have stool characteristics more similar to those of breastfed infants, suggesting
a positive effect of prebiotics on the digestive system. Preliminary evidence also suggests that
prebiotics may positively affect mineral absorption and the infant immune system.
The neonatal period is a critical period of development, when potentially long-term effects of
manipulation of the intestinal microbiota could result.141 While the clinical data are promising,
more studies are needed to understand the conditions under which prebiotic formula ingredients
can positively influence infant growth and development. Mead Johnson continues to evaluate the
science supporting the use of prebiotic ingredients in infant formula with the goal of providing
state-of-the-science nutrition to formula-fed infants.
26
27
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